Next-generation sequencing (NGS) has allowed for the rapid characterization of genomes, exomes and transcriptomes. Such advances have been applied to personalized oncology, represent a promising approach for identifying therapeutic options for cancer patients who do not respond to standard treatments, and are key to improving our understanding of tumorigenesis. However, although the cost of performing whole genome sequencing (WGS) has decreased in recent years, it is more costly compared to exome and RNA sequencing (RNAseq) when sequencing to 30× coverage. Owing to this caveat and the existing utility of using deep exome sequencing to identify potentially targetable small somatic events in cancer genomes, the need for identifying an alternative WGS strategy for identifying breakpoints, which characterize structural variants and copy number changes, is clear.
One option for evaluating larger regions in whole genome data using sequencing by synthesis (SBS) technology is the use of Illumina's mate pair library preparation protocol. The standard protocol requires 10 μg of genomic DNA and supports the evaluation of regions spanning up to approximately 2-5 kb. However, owing to the limited amount of DNA that is typically available from tumor biopsies, this approach is not a viable option for sequencing. Illumina also recently released a new Nextera Mate Pair Sample Preparation Kit that requires 1-4 μg of genomic DNA. However, this approach retains transposome-mediated fragmentation that results in an enzymatic footprint that requires trimming of sequencing data, and still requires circularization and biotin pull-down, and thus decreases the ease of library preparation. An alternative user-friendly strategy that requires lower inputs, that does not require post-sequencing trimming and that allows for increased physical coverage and analysis of regions greater than that accomplished by short insert (SI) sequencing is thus needed.